1. Field of the Invention
The present invention relates generally to the fields of molecular biology and medicine. More particularly, it concerns methods and compositions for identifying candidate therapeutic and preventative agents and for treating or preventing diseases, disorders and conditions involving trinucleotide repeats.
2. Description of Related Art
Myotonic dystrophy, the most common adult-onset muscular dystrophy in humans, is characterized by the severe dysfunction of skeletal and smooth muscle structure and function (Harper, 2001; Groenen and Wieringa, 1998; Mankodi and Thornton, 2002; Amack and Mahadevan, 2004). Delayed muscle differentiation and hypotonia are the predominant features in congenital myotonic dystrophy type 1 (DM1) whereas myotonia, atrophy and weakness of skeletal muscle are the features in adult onset DM1 (Harper, 2001; Sarnat and Silbert, 1976; Farkas-Bargeton et al., 1988; Sahgal et al., 1983). Several dystrophic changes of smooth muscles are also common in DM1 (Lenard et al., 1977; Brunner et al., 1992; Ronnblom et al., 1999; Ronnblom et al., 2002). DM1 genetic mutation is the expansion of a CTG repeat in the 3′ un-translated region of DMPK on chromosome 19q13.3 and DM2 (myotonic dystrophy type 2) genetic mutation is the expansion of a CCTG repeat in the first intron of ZNF9 located on chromosome 3q21.3 (Fu et al, 1992; Brook et al., 1992; Mahadevan et al., 1992; Liquori et al., 2001.
In DM1, the length of the CTG repeat shows a strong correlation with the complexity and severity of disease phenotypes; short to moderate expansion causes milder muscle defects whereas expansions larger than 3 kb cause more severe dystrophic muscle defects (Harper, 2001; Monckton and Ashizawa, 2004; Nagamitsu and Ashizawa, 2002). In contrast, DM2 phenotypes are more subtle and DM2 patients do not develop congenital muscle defects in spite of very large repeat expansion (Day et al., 1999; Day et al., 2003). In DM1, transcription of the mutant DMPK allele produces mRNA encoding expanded CUG sequences; in DM2, the transcription of ZNF9 produces mRNA that carries expanded CCUG sequences. The CUG and CCUG RNA cause cellular toxicity, but the mechanism by which expanded CUG and CCUG RNA cause such cellular toxicity has not been elucidated completely (Mankodi et al., 2003; Kanadia et al., 2003; Jiang et al., 2004; Day and Ranum, 2005; Ranum L P, Day, 2004). Transgenic mice expressing short stretches of CUG repeats (approximately 250 repeats) in skeletal muscle develop myotonia and several structural defects reminiscent of DM1 (Mankodi et al., 2000) Sequestration of muscle-blind proteins by expanded CUG RNA was found to be the pathogenic mechanism in the muscle defects found in the CUG transgenic mice (Mankodi et al., 2002). Subsequent studies with muscle-blind (Mbnl1) knockout mice confirmed that the loss of muscle-blind function is indeed one of the important mechanistic steps required to inflict muscle defects in DM1 (Kanadia et al., 2003). Importantly, neither Mbnl1 knockout mice nor the available CUG transgenic mice develop dystrophic muscle defects, the hallmark clinical features of adult onset and congenital DM1 (Kanadia et al., 2003; Mankodi et al., 2000). These complex muscle defects include severe muscle weakness, wasting, degeneration of muscle fibers and developmental abnormalities of skeletal muscle fibers.
Therefore, there remains a need to recapitulate the physiological context of DM1 in order to identify key proteins involved in DM1, and thus allow for the identification and development of appropriate therapeutic and preventative agents for the disease.
Moreover, DM1 represents an example of a disease associated with repeat sequences in nucleic acid molecules. Many other diseases, as well as conditions and disorders, do not have cures, cannot be prevented, or are not effectively treated. These would benefit from compositions and methods that would elucidate the mechanisms behind their pathologies so as to lead to the identification and development of candidate drugs. Consequently, there is a need for methods and compositions that would provide information regarding other diseases, conditions, and disorders associated with repeat sequences.